The invention relates to fluorescent dyes (fluorophores) based on polymethines for use in optical measurement and detection procedures, in particular those employing fluorescence. Typical applications exploit the reaction of dye-labelled antigens, antibodies or DNA segments with the respective complementary species. With such methods it is possible to measure, e.g., enzyme kinetics, receptor-ligand interactions and the kinetics of nucleic-acid hybridization. Furthermore, the claimed fluorophores are of interest for the pharmacological characterization of receptors or agents.
Possible uses associated therewith exist, for example, in medicine and pharmacology, in the biological and materials sciences, in environmental monitoring and the detection of organic and inorganic microsamples present in natural surroundings and in technology.
Whereas cyanines with a Stokes shift of 20-40 nm have long been known as fluorescent markers (Cy3, Cy5, U.S. Pat. No. 5,627,027), as yet there are only a few fluorophores with a large Stokes shift. Typical examples of these are the markers derived from the laser dye DCM, with absorption maxima at 481 nm and emission maxima at 644 nm, which are claimed in the patent U.S. Pat. No. 4,886,744.
Different stilbenes with large Stoke's shift have been previously described by F. Lehmann et al. in “Dyes and Pigments”, Elsevier Applied Science Publishers, vol. 29, no. 1, 1995, pages 85-94 as well as by G. J. Ashwell et al. in the “Journal of Materials Chemistry”, Cambridge, vol. 11, no. 5, 2001, pages 1345-50. Hydroxy substituted-stilbenes with a lactone bridging have been published by R. M. Abd El-Aal et al. in the “Journal of the Chinese Chemical Society”, Taipei, vol. 47, no. 2, 2000, pages 389-95. Further to be taken into consideration are the different stilbenes published by Czerney et al. in “Sensors and Actuators B”, Elsevier, Lausanne, vol. 39, no. 1-3, 1997, pages 395-400 and in U.S. Pat. No. 6,096,794.
Common to all compounds is that they do not have any reactive group which permits covalent binding, for example, biomolecules. A further disadvantage is that the solubility in water or aqueous media for applications in bioanalysis is not sufficient.
Whereas the compounds disclosed in the Japanese unexamined patent specification JP 07-234 504 contain an —SO3-group, they have a completely different structure than the compounds I to III according to the invention. They are distinguished from the latter in that they are not suited for applications in bioanalysis and diagnosis because they lack a function which enables a covalent binding on biomolecules.